期刊文献

Highly efficient prime editing by introducing same-sense mutations in pegRNA or stabilizing its structure 收藏

通过在Pegrna中引入相同的突变或稳定其结构，高效的Prime编辑

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作者

作者单位

[1]Department of Ophthalmology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China;[2]Gene Editing Center, School of Life Science and Technology, ShanghaiTech University, Shanghai, China;[3]CAS Key Laboratory of Computational Biology, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China;[4]Shanghai Clinical Research and Trial Center, Shanghai, China;[5]Center for Molecular Medicine, Children’s Hospital, Fudan University and Shanghai Key Laboratory of Medical Epigenetics, International Laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, China

关键词

CRISPR-Cas9 genome editing;Genetic engineering;Science;Humanities;Social Sciences;multidisciplinary

关键词译文

CRISPR-CAS9基因组编辑；基因工程；科学；人文科学；社会科学；多学科

发布日期

29 March 2022

页码

1-9

DOI

10.1038/s41467-022-29339-9

来源信息

Scientific reports ISSN：2045-2322, 2022年, 1-9页

摘要

Prime editor (PE), which is developed by combining Cas9 nickase and an engineered reverse transcriptase, can mediate all twelve types of base substitutions and small insertions or deletions in living cells but its efficiency remains low. Here, we develop spegRNA by introducing same-sense mutations at proper positions in the reverse-transcription template of pegRNA to increase PE’s base-editing efficiency up-to 4,976-fold (on-average 353-fold). We also develop apegRNA by altering the pegRNA secondary structure to increase PE’s indel-editing efficiency up-to 10.6-fold (on-average 2.77-fold). The spegRNA and apegRNA can be combined to further enhance editing efficiency. When spegRNA and apegRNA are used in PE3 and PE5 systems, the efficiencies of sPE3, aPE3, sPE5 and aPE5 systems are all enhanced significantly. The strategies developed in this study realize highly efficient prime editing at certain previously uneditable sites.

摘要译文

Prime Editor（PE）是通过将Cas9 Nickase和一种工程化逆转录酶结合而开发的，可以介导所有十二种类型的基础取代，而在活细胞中进行小插入或缺失，但其效率仍然很低。在这里，我们通过在Pegrna的反向转录模板中引入相同的突变来发展Spegrna，以提高PE的基础编辑效率，最高为4,976倍（353倍）。我们还通过改变PEGRNA二级结构来发展Apegrna，以提高PE的ideel编辑效率至10.6倍（平均2.77倍）。Spegrna和Apegrna可以组合以进一步提高编辑效率。当Spegrna和Apegrna用于PE3和PE5系统中时，SPE3，APE3，SPE5和APE5系统的效率都得到了显着增强。这项研究中制定的策略在某些以前无法编辑的站点上实现了高效的质量编辑。

Li; Xiaosa[1];Zhou; Lina[2];Gao; Bao-Qing[3];Li; Guangye[2];Wang; Xiao[2];Wang; Ying[3];Wei; Jia[3];Han; Wenyan[2];Wang; Zixian[2];Li; Jifang[2];Gao; Runze[2];Zhu; Junjie[2];Xu; Wenchao[2];Wu; Jing[2];Yang; Bei[4];Sun; Xiaodong[1];Yang; Li[5];Chen; Jia[2]. Highly efficient prime editing by introducing same-sense mutations in pegRNA or stabilizing its structure[J]. Scientific reports, 2022: 1-9